PROJECT SUMMARY/ABSTRACT Dengue virus cause millions of infections and thousands of deaths annually worldwide. Antibody-dependent enhancement (ADE) is thought to trigger most cases of severe DV infection and is a key consideration in DV vaccine development. ADE occurs when antibodies elicited by infection with a heterologous DV strain form immune complexes (ICs) that allow virus entry into endosomes of myeloid cells via Fc? receptors (Fc?Rs). Once within endosomes, successful infection requires a pH-dependent conformational change in the DV envelope resulting in fusion with late endosomal membranes. Notably, endosomes of myeloid cells reliably express Fc neonatal receptors (FcRns), which are distinct from Fc?Rs and which engage IgG at acidic pH and release it at neutral pH. Thus, in the case of DV ICs, there is an intersection within endosomes between envelope fusion and engagement of the ICs with FcRn. Until now, however, the effect of such engagement on ADE has not been studied. Our preliminary data point strongly toward an important role for FcRn in DV ADE, such that abrogating IC binding to FcRn reduces ADE, whereas increasing IC-FcRn binding increases ADE. In the proposed research, we will use a novel mouse model to determine if, consistent with our in vitro data, FcRn engagement with DV ICs increases ADE and the severity of infection outcomes. We will also use in vitro models to explore potential mechanisms that underlie FcRn involvement in ADE. We will test the following hypotheses: 1. FcRn engagement by antibodies bound to DV increases the likelihood of ADE and of severe infection outcomes; and 2. After internalization of DV ICs via Fc?Rs, FcRn binding results in retaining infectious ICs in endosomes, thus increasing the likelihood of successful infection. To test these hypotheses, we will accomplish two specific aims: 1. Determine the impact of human mAbs, with and without Fc mutations that alter FcRn binding, on the course of DV infection in interferon receptor-deficient (Ifnar1-/-) mice expressing human FcRn. In addition, we will compare ADE in Ifnar1-/- mice with and without FcRn expression; 2. Determine the intracellular fate of DV ICs in myeloid cells. The trajectory of DV ICs and the timing and site of fusion between the virus envelope and endosomal membranes will be tracked by fluorescence microscopy. This research may lead to new therapeutic strategies for treating severe dengue virus infections, such as the use of FcRn-blocking antibodies or drugs, and to novel insights into DV vaccine development. More broadly, if FcRn is mechanistically involved in DV replication in the presence of antibody, it is very plausible that a similar mechanism might be involved in the replication of any of the several viruses whose life cycle requires the acidic milieu of the endosome.